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Cytoviva Hyperspectral Microscopy – Visible Near Infrared (VNIR) and Short Wave Infrared (SWIR)

CytoViva’s Hyperspectral Imaging technology was specifically designed to provide quantitative spectral analysis of nanoscale materials imaged with the patented CytoViva darkfield-based microscope system or with other microscopy modalities

This can include spectral analysis of both biological and materials-based nanoscale samples, which may be isolated or integrated in cells, tissue or other materials-based matrices. CytoViva provides hyperspectral imaging in the visible near-infrared (VNIR 400nm-1,000nm) wavelength ranges as well as in the short wave infrared (SWIR 900nm-1,700nm) wavelength ranges.

In addition, CytoViva provides hyperspectral imaging in both wavelength ranges for macro-level samples that do not require microscopy level analysis.

Hyperspectral images appear very similar to a traditional optical image with one important difference. When observed via image analysis software, each pixel of a hyperspectral image provides the complete spectral response of that pixel’s spatial area within the VNIR or SWIR spectral range. At 100x magnification, a hyperspectral image may contain as many as 700,000 pixels as small as 128nm each. This spectral data is recorded at approximately 2nm of spectral resolution in the VNIR range, enabling minute spectral differences to be measured from pixel to pixel within the image.

Using CytoViva’s proprietary hyperspectral image analysis software, it is possible to identify, locate and map nano-scale materials within a sample based on the unique spectral response of this material. This can be accomplished without any special staining or tagging of the target material, as is often required with traditional microscopy techniques. In addition, when functional groups are added onto or into a nano-scale sample, they can often be identified, as this change in the sample’s surface chemistry will be spectrally detected.

The primary components of CytoViva’s Hyperspectral Imaging technology include a transmission diffraction grating spectrograph and an integrated camera. These components are mounted onto the camera (C) mount of the microscope. A hyperspectral image is created in a line scan or “pushbroom” fashion, by moving the sample across the field of view of the microscope and spectrograph via an automated translational microscope stage. Typically these spectral images are created in seconds or minutes, depending on the required exposure. The hyperspectral imaging hardware is integrated with customized image analysis software. This proprietary software provides the ability to compare spectra within a sample image or between images. It can also build a spectral library of unique samples elements. Using this spectral library, these sample elements can then be mapped in subsequent samples.

To learn more about the research applications supported by CytoViva’s Hyperspectral Microscopy System, please click on the hyperspectral applications page above.

The CytoViva® Microscope System – Advanced Darkfield Illumination System

Advanced Darkfield Illumination System

The CytoViva advanced darkfield illumination system replaces the standard microscope condenser. The specialized illuminator focuses fixed-geometry, highly collimated light at oblique angles on the sample. This serves to dramatically improve contrast and signal-to-noise ratio which allows for optimized resolving power and detection capability of non-fluorescing samples.

The CytoViva 3D Enhanced Darkfield Imaging System

The CytoViva 3D Enhanced Darkfield Imaging System provides a method for locating non-labeled nanostructures (particles, tubes, etc.) in a variety of translucent matrices (cells, tissue, organisms). This technique leverages the high signal-to-noise optical performance of the patented CytoViva Enhanced Darkfield Microscope Technology in combination with patent-pending deconvolution and particle location routines to provide users with a three dimensional optical model of their sample. Most importantly, this technique does not require the use of fluorescent labels on the nanoparticles to obtain an image of the particles, thus removing the potentially negative influence of these labels on the sample.

The 3D image is created by acquiring and storing a “stack” of equally spaced two dimensional images using the CytoViva Enhanced Darkfield Microscope Technology, a piezo-driven Z-axis stage and CytoViva’s proprietary image acquisition and control software.  This stack of images is then processed using CytoViva 3D Image Analysis ImageJ plug-ins to locate the subject particles, as well as, deconvolve the surrounding cells, tissue or other translucent matrix.  The resulting optical image is then viewed in an ImageJ 3D viewer.  The use of ImageJ also allows further processing of the data in an environment familiar to many users.

The images above  illustrate three examples of  the 3D capability.  Figure 1 is a 3D deconvolution  of nanoparticles  (NPs) in a cell with the NPs in red, the nuclear structure in blue and the  cell structure in gray. Figure 2  is a 3D deconvolution of a carbon nanotube piercing a tissue section.

The CytoViva 3D Enhanced Darkfield Imaging System is designed to facilitate research in a variety of applications in nanobiology (targeted drug delivery and pathogen detection), nanotoxicology (nanoparticles and carbon nanotubes in tissue), and others where it is useful to understand the location of nanostructures in a three dimensional space.  This capability is fully compatible with CytoViva’s Hyperspectral Imaging system and can be operated on the same microscope platform, thus allowing existing systems to be easily upgraded to include this capability.

3D Videos:

Unlabeled Au Nanoparticles in a Cell

Unlabeled Carbon Nanotubes Embedded in Tissue

Fiber Matrix